La maladie d'Alzheimer (MA) a un impact socio-économique majeur. Le diagnostic n’est certain qu’à l'autopsie. Il n’existe pas de biomarqueur assurant un diagnostic précoce in vivo. Les quatre traitements actuellement disponibles sont symptomatiques. Les autres traitements qui ont fait l’objet d’essais de recherche clinique, n’ont pas démontré de modification de l’évolution de la maladie. Les hypothèses sous-jacentes à cet échec sont soit l’inefficacité du traitement, soit un manque d'homogénéité de la population étudiée (diagnostic différentiel parfois très compliqué), soit l’absence de biomarqueurs fiables in vivo en mesure de détecter une modification. Dans ce contexte, il est essentiel d'identifier un biomarqueur in vivo qui permette d’établir le diagnostic différentiel entre la MA et les autres démences et d’évaluer l'efficacité des traitements. Ce travail vise à caractériser in vitro, ex vivo et in vivo le 3 - (3 - (3-fluoropropylthio) -1,2,5-thiadiazole-4-yl) -1,2,5,6-tétrahydro-1-méthylpyridine [FP-TZTP], chez les rongeurs pour évaluer son intérêt en tant que biomarqueur potentiel pour la maladie d'Alzheimer. Après avoir établi que l’expression du sous type M2 des récepteurs muscariniques était modifiée précocement dans la MA, nous avons mis en évidence une distribution du radiotraceur fluoré FP-TZTP spécifique de M2 chez la souris génétiquement modifiée ‘knock out’. Afin de mieux caractériser ce radiotraceur, nous avons effectué des études de culture cellulaire in vitro ainsi que des études ex-vivo sur du tissu cérébral pour comprendre la spécificité du [18F]FP-TZTP (Résultats #1). Les résultats ex-vivo nous ont encouragés à réaliser les études in vivo. Nous avons choisi la tomographie par émission de positons (TEP) qui permet l’étude in vivo et non invasive de la biodistribution du [18F]FP-TZTP chez le rat, en utilisant le scanner pour animaux de petite taille (ATLAS) développé par « The National Institutes of Health , Bethesda, MD, USA» (Résultats #4). Pour cela, nous avons dans un premier temps testé l'ATLAS au moyen de deux études métaboliques chez le rat, avec un traceur couramment utilisé, le [18F]fluorodéoxyglucose ([18F]FDG) (Résultats #2; Résultats #3). Nos études suggèrent que le [18F]FP-TZTP est un traceur approprié pour la quantification en TEP des récepteurs muscariniques de type M2, utile pour le diagnostic précoce de la MA. / Alzheimer’s disease (AD) has an increasingly critical impact on society from the socio-economic point of view in addition to being very burdensome for the patients themselves, their relatives and friends. Diagnosis of certitude is only at post mortem and no single biomarker has yet been found to be accurate for early in vivo diagnosis. The current available treatments are only symptomatic. The few treatments under research trials have failed to demonstrate a disease modification for either lack of actual treatment efficacy or for lack of population homogeneity and for lack of reliable in vivo biomarkers able to detect a modification. In this context, it is both urgent and necessary to identify an in vivo biomarker that enables i) the differential diagnosis of AD among other dementias and ii) the assessment of treatment efficacy as a follow up in AD patients, is clearly very noticeable. This work aims to characterize the 3-(3-(3-fluoropropylthio)-1,2,5-thiadiazol-4-yl)-1,2,5,6-tetrahydro-1-methylpyridine [FP-TZTP], by use of in vitro, ex–vivo and in vivo methods in rodents, to assess whether it is a suitable biomarker for Alzheimer’s disease. An impairment of the M2 subtype of the muscarinic receptors was noticed in AD patients and clear evidences of M2 selectivity in knock out mice previously injected with the fluorinated radiotracer FP-TZTP was observed. To further characterize such M2 selectivity, we performed in vitro cell culture and ex-vivo tissue dipping studies (Results #1). Encouraged by the ex-vivo results, we went on to the in vivo world. We elected the non-invasive nuclear medicine imaging technique Positron Emission Tomography (PET) to assess the biodistribution of the [18F]FP-TZTP in rats by use of the Advanced Technology Laboratory Animal Scanner (ATLAS) developed at the National Institutes of Health, Bethesda, MD, USA (Results #4). We had first assessed the ATLAS as a legitimate tool by use of a commonly used and well known radiotracer, the [18F]fluorodeoxyglucose ([18F]FDG) (Results #2 and #3). Our studies suggest that [18F]FP-TZTP may be a biomarker for AD as it is a suitable tracer for in vivo quantification of the M2 receptors.

Elevated sympathetic nervous system (SNS) tone contributes to excess cardiac mortality associated with type 2 diabetes mellitus (T2DM). Chronic SNS stimulation has detrimental effects to the heart, in particular, with its cell signaling abilities. (R)-[11C]Rolipram small animal positron emission tomography (PET), an noninvasive nuclear imaging modality, was used to assess phosphodiesterase-4 (PDE4) alterations in a high fat diet (HFD), streptozotocin (STZ) induced model of hyperglycemia in rats. Prior to investigation in the animal model, characterization of (R)-[11C]rolipram small animal PET was completed. (R)-[11C]Rolipram binds specifically to PDE4 in the rat heart demonstrated by competitive blockade with (R)-rolipram with the PDE4 enzyme susceptible to saturation with increasing injected masses of unlabeled rolipram. (R)-[11C]Rolipram cardiac retention was elevated by acute norepinephrine stimulation via desipramine pharmacologic challenge. Quantitative (R)-[11C]rolipram PET was highly reproducible in the heart and presents an ideal avenue to investigate PDE4 alterations. (R)-[11C]rolipram small animal PET did not reveal changes in PDE4 expression and activity in STZ-treated hyperglycemic animals compared to STZ-treated euglycemic and control groups. In vitro measures of PDE4 enzyme expression and activity, with or without desipramine, were also not altered between treatment groups. Although (R)-[11C]rolipram small animal PET does not reveal PDE4 alterations in this animal model of diabetes, its utility to assess PDE4 alterations in other over active SNS pathologies, such as heart failure and obesity, remains.

Elevated sympathetic nervous system (SNS) tone contributes to excess cardiac mortality associated with type 2 diabetes mellitus (T2DM). Chronic SNS stimulation has detrimental effects to the heart, in particular, with its cell signaling abilities. (R)-[11C]Rolipram small animal positron emission tomography (PET), an noninvasive nuclear imaging modality, was used to assess phosphodiesterase-4 (PDE4) alterations in a high fat diet (HFD), streptozotocin (STZ) induced model of hyperglycemia in rats. Prior to investigation in the animal model, characterization of (R)-[11C]rolipram small animal PET was completed. (R)-[11C]Rolipram binds specifically to PDE4 in the rat heart demonstrated by competitive blockade with (R)-rolipram with the PDE4 enzyme susceptible to saturation with increasing injected masses of unlabeled rolipram. (R)-[11C]Rolipram cardiac retention was elevated by acute norepinephrine stimulation via desipramine pharmacologic challenge. Quantitative (R)-[11C]rolipram PET was highly reproducible in the heart and presents an ideal avenue to investigate PDE4 alterations. (R)-[11C]rolipram small animal PET did not reveal changes in PDE4 expression and activity in STZ-treated hyperglycemic animals compared to STZ-treated euglycemic and control groups. In vitro measures of PDE4 enzyme expression and activity, with or without desipramine, were also not altered between treatment groups. Although (R)-[11C]rolipram small animal PET does not reveal PDE4 alterations in this animal model of diabetes, its utility to assess PDE4 alterations in other over active SNS pathologies, such as heart failure and obesity, remains.

Small animal positron emission tomography (PET) image analysis can be particularly challenging with heavily-lesioned animal disease models with limited tracer uptake such as the 6-hydroxydopamine (OHDA) lesioned rat model of Parkinson's disease. Methodology-related variations in measured values of 10% or 15% can obscure meaningful biological differences, so accurate analysis methods are essential. However, placing regions of interest (ROIs) on these images without additional guidance is unreliable, and can lead to significant errors in results. To address this problem, this work develops a partly atlas-guided method place ROIs on structures that lack specific binding with presynaptic dopaminergic tracers. The method is tested by correlation of PET binding potential (BP) with autoradiographic binding measurements, and with repeated PET scans of the same subjects, both with the presynaptic tracer ¹¹C-dihydrotetrabenazine (DTBZ). The method is found to produce reliable results. When directly comparing PET images of the same subject to detect changes, it is essential to minimize variations due to analysis method. To this end, a masking method for automated image registration (AIR) of PET images with dopaminergic tracer rat images is developed. Coregistration with AIR and separate ROI placement are compared and tested with repeated scans of the same rat with DTBZ, and are found to be equivalent. Kinetic modelling algorithms may also introduce bias or scatter to binding potentials (BP) calculated from TACs or in parametric images. To determine the optimal method for this step, algorithms for dopaminergic tracers are compared for small animal DTBZ, ¹¹C-methylphenidate (MP), and ¹¹C-raclopride (Rac) data. Among the tested methods is a new variant of the Logan graphical kinetic modelling method, developed in this work, that issignificantly less biased by target tissue TAC noise than the standard Logan approach. The modified graphical method is further compared with the Logan graphical algorithms with added-noise simulations. The simplified reference tissue model (SRTM) is found to have the best method for ROI TAC data, while the modified graphical algorithm may be preferred when generating parametric images.

Non-invasive dynamic optical imaging of small animals requires the development of a novel fluorescence imaging modality. Herein, fluorescence imaging is demonstrated with sub-second camera integration times using agents specifically targeted to disease markers, enabling rapid detection of cancerous regions. The continuous-wave fluorescence imaging acquires data with an intensified or an electronmultiplying charge-coupled device. The work presented in this dissertation (i) assessed dose-dependent uptake using dynamic fluorescence imaging and pharmacokinetic (PK) models, (ii) evaluated disease marker availability in two different xenograft tumors, (iii) compared the impact of autofluorescence in fluorescence imaging of near-infrared (NIR) vs. red light excitable fluorescent contrast agents, (iv) demonstrated dual-wavelength fluorescence imaging of angiogenic vessels and lymphatics associated with a xenograft tumor model, and (v) examined dynamic multi-wavelength, whole-body fluorescence imaging with two different fluorescent contrast agents. PK analysis showed that the uptake of Cy5.5-c(KRGDf) in xenograft tumor regions linearly increased with doses of Cy5.5-c(KRGDf) up to 1.5 nmol/mouse. Above 1.5 nmol/mouse, the uptake did not increase with doses, suggesting receptor saturation. Target to background ratio (TBR) and PK analysis for two different tumor cell lines showed that while Kaposi’s sarcoma (KS1767) exhibited early and rapid uptake of Cy5.5-c(KRGDf), human melanoma tumors (M21) had non-significant TBR differences and early uptake rates similar to the contralateral normal tissue regions. The differences may be due to different compartment location of the target. A comparison of fluorescence imaging with NIR vs. red light excitable fluorescent dyes demonstrates that NIR dyes are associated with less background signal, enabling rapid tumor detection. In contrast, animals injected with red light excitable fluorescent dyes showed high autofluorescence. Dual-wavelength fluorescence images were acquired using a targeted 111In- DTPA-K(IRDye800)-c(KRGDf) to selectively detect tumor angiogenesis and an untargeted Cy5.5 to image lymphatics. After acquiring the experimental data, fluorescence image-guided surgery was performed. Dynamic, multi-wavelength fluorescence imaging was accomplished using a liquid crystal tunable filter (LCTF). Excitation light was used for reflectance images with a LCTF transmitting a shorter wavelength than the peak in the excitation light spectrum. Therefore, images can be dynamically acquired alternating frame by frame between emission and excitation light, which should enable image-guided surgery.

Small animal positron emission tomography (PET) image analysis can be particularly challenging with heavily-lesioned animal disease models with limited tracer uptake such as the 6-hydroxydopamine (OHDA) lesioned rat model of Parkinson's disease. Methodology-related variations in measured values of 10% or 15% can obscure meaningful biological differences, so accurate analysis methods are essential. However, placing regions of interest (ROIs) on these images without additional guidance is unreliable, and can lead to significant errors in results. To address this problem, this work develops a partly atlas-guided method place ROIs on structures that lack specific binding with presynaptic dopaminergic tracers. The method is tested by correlation of PET binding potential (BP) with autoradiographic binding measurements, and with repeated PET scans of the same subjects, both with the presynaptic tracer ¹¹C-dihydrotetrabenazine (DTBZ). The method is found to produce reliable results. When directly comparing PET images of the same subject to detect changes, it is essential to minimize variations due to analysis method. To this end, a masking method for automated image registration (AIR) of PET images with dopaminergic tracer rat images is developed. Coregistration with AIR and separate ROI placement are compared and tested with repeated scans of the same rat with DTBZ, and are found to be equivalent. Kinetic modelling algorithms may also introduce bias or scatter to binding potentials (BP) calculated from TACs or in parametric images. To determine the optimal method for this step, algorithms for dopaminergic tracers are compared for small animal DTBZ, ¹¹C-methylphenidate (MP), and ¹¹C-raclopride (Rac) data. Among the tested methods is a new variant of the Logan graphical kinetic modelling method, developed in this work, that issignificantly less biased by target tissue TAC noise than the standard Logan approach. The modified graphical method is further compared with the Logan graphical algorithms with added-noise simulations. The simplified reference tissue model (SRTM) is found to have the best method for ROI TAC data, while the modified graphical algorithm may be preferred when generating parametric images.

This thesis reports on a novel use of the Medipix All Resolution System (MARS) Spectral CT scanner as a platform of small animal irradiation. Irradiation from the x-ray source in the scanner was used to inhibit the growth and recovery of neurons in rats. The goal is to extrapolate the relationship between stem cells and memory and functional behaviours. Specific developments were carried out prior to the irradiation of live rats. A live rat holder was designed and built to provide a setup stable enough for fixing the position of the rats head. This was achieved by integrating a three piece bar system into the holder to hold the head from both ears and nose. To quantify the absorbed dose, x-ray exposures were measured using a calibrated ion chamber and were used to generate a depth dose curve with sheets of Perspex layers and radiochromic films. This curve is presumed to simulate the dose inside the rat‘s head. To target a sub region of interest within the rat‘s brain, specific anatomical landmarks were investigated for the design of the lead collimators. The position of the sub volume, the hippocampus, was located through a combination of anatomical landmarks and x-ray transmission images of the rat‘s head. Bregma and the interaural line were used to numerically plot out the co-ordinates of the dorsal and ventral hippocampus which was then translated onto the lead collimators. The x-ray transmission images of euthanized rats were used as a guide to locate the dorsal and ventral hippocampuses. Bregma and the interaural line were the main anatomical landmarks which were used for the design of the lead collimators to be placed around the head. Three pilot rats were irradiated with the designed holder and collimators. The point dose to the hippocampus was calculated using the simulated depth dose curve. Post irradiation status of the neurogenesis was assessed three weeks after the treatment. However only one of the three rats showed a significant reduction in the number of neurons in the hippocampus emphasizing the room for more improvement in the physical setup of the irradiation.

Elevated sympathetic nervous system (SNS) tone contributes to excess cardiac mortality associated with type 2 diabetes mellitus (T2DM). Chronic SNS stimulation has detrimental effects to the heart, in particular, with its cell signaling abilities. (R)-[11C]Rolipram small animal positron emission tomography (PET), an noninvasive nuclear imaging modality, was used to assess phosphodiesterase-4 (PDE4) alterations in a high fat diet (HFD), streptozotocin (STZ) induced model of hyperglycemia in rats. Prior to investigation in the animal model, characterization of (R)-[11C]rolipram small animal PET was completed. (R)-[11C]Rolipram binds specifically to PDE4 in the rat heart demonstrated by competitive blockade with (R)-rolipram with the PDE4 enzyme susceptible to saturation with increasing injected masses of unlabeled rolipram. (R)-[11C]Rolipram cardiac retention was elevated by acute norepinephrine stimulation via desipramine pharmacologic challenge. Quantitative (R)-[11C]rolipram PET was highly reproducible in the heart and presents an ideal avenue to investigate PDE4 alterations. (R)-[11C]rolipram small animal PET did not reveal changes in PDE4 expression and activity in STZ-treated hyperglycemic animals compared to STZ-treated euglycemic and control groups. In vitro measures of PDE4 enzyme expression and activity, with or without desipramine, were also not altered between treatment groups. Although (R)-[11C]rolipram small animal PET does not reveal PDE4 alterations in this animal model of diabetes, its utility to assess PDE4 alterations in other over active SNS pathologies, such as heart failure and obesity, remains.

Small animal positron emission tomography (PET) image analysis can be particularly challenging with heavily-lesioned animal disease models with limited tracer uptake such as the 6-hydroxydopamine (OHDA) lesioned rat model of Parkinson's disease. Methodology-related variations in measured values of 10% or 15% can obscure meaningful biological differences, so accurate analysis methods are essential. However, placing regions of interest (ROIs) on these images without additional guidance is unreliable, and can lead to significant errors in results. To address this problem, this work develops a partly atlas-guided method place ROIs on structures that lack specific binding with presynaptic dopaminergic tracers. The method is tested by correlation of PET binding potential (BP) with autoradiographic binding measurements, and with repeated PET scans of the same subjects, both with the presynaptic tracer ¹¹C-dihydrotetrabenazine (DTBZ). The method is found to produce reliable results. When directly comparing PET images of the same subject to detect changes, it is essential to minimize variations due to analysis method. To this end, a masking method for automated image registration (AIR) of PET images with dopaminergic tracer rat images is developed. Coregistration with AIR and separate ROI placement are compared and tested with repeated scans of the same rat with DTBZ, and are found to be equivalent. Kinetic modelling algorithms may also introduce bias or scatter to binding potentials (BP) calculated from TACs or in parametric images. To determine the optimal method for this step, algorithms for dopaminergic tracers are compared for small animal DTBZ, ¹¹C-methylphenidate (MP), and ¹¹C-raclopride (Rac) data. Among the tested methods is a new variant of the Logan graphical kinetic modelling method, developed in this work, that issignificantly less biased by target tissue TAC noise than the standard Logan approach. The modified graphical method is further compared with the Logan graphical algorithms with added-noise simulations. The simplified reference tissue model (SRTM) is found to have the best method for ROI TAC data, while the modified graphical algorithm may be preferred when generating parametric images. / Science, Faculty of / Physics and Astronomy, Department of / Graduate

Elevated sympathetic nervous system (SNS) tone contributes to excess cardiac mortality associated with type 2 diabetes mellitus (T2DM). Chronic SNS stimulation has detrimental effects to the heart, in particular, with its cell signaling abilities. (R)-[11C]Rolipram small animal positron emission tomography (PET), an noninvasive nuclear imaging modality, was used to assess phosphodiesterase-4 (PDE4) alterations in a high fat diet (HFD), streptozotocin (STZ) induced model of hyperglycemia in rats. Prior to investigation in the animal model, characterization of (R)-[11C]rolipram small animal PET was completed. (R)-[11C]Rolipram binds specifically to PDE4 in the rat heart demonstrated by competitive blockade with (R)-rolipram with the PDE4 enzyme susceptible to saturation with increasing injected masses of unlabeled rolipram. (R)-[11C]Rolipram cardiac retention was elevated by acute norepinephrine stimulation via desipramine pharmacologic challenge. Quantitative (R)-[11C]rolipram PET was highly reproducible in the heart and presents an ideal avenue to investigate PDE4 alterations. (R)-[11C]rolipram small animal PET did not reveal changes in PDE4 expression and activity in STZ-treated hyperglycemic animals compared to STZ-treated euglycemic and control groups. In vitro measures of PDE4 enzyme expression and activity, with or without desipramine, were also not altered between treatment groups. Although (R)-[11C]rolipram small animal PET does not reveal PDE4 alterations in this animal model of diabetes, its utility to assess PDE4 alterations in other over active SNS pathologies, such as heart failure and obesity, remains.